Bradbury [2045] proposes
using artificial enzymes for nanoscale parts assembly. After noting Merkle’s
[2017] suggestion that the positional control
of nanoscale building blocks would allow the extension of the normal reactions
found in biological systems to include free-radical chemistry and more recent
descriptions of similar reactive chemistries being employed by enzymes [2046],
and related informal suggestions for artificial mechanoenzymes by Freitas [2047]
and “mechalysts” by Craver [2048],
Bradbury [2045] envisioned the engineering
of artificial multifunctional proteins called “robozymes” having
the following properties: (1) unfolded, the robozyme grabs onto molecular building
blocks [2088-2090],
carefully keeping them separate from each other to avoid nonspecific reactions;
(2) using specific enzyme catalytic sites near the bound building blocks, it
“activates” the molecules (perhaps producing one or more free radicals);
(3) induced folding brings the building blocks into relatively precise alignment
allowing the desired chemical reaction(s) to occur; and (4) the protein is induced
to unfold, releasing the final product. With protein folding forces in the tens
to hundreds of pN, such enzymes could also provide a means for threading one
molecule through another molecule, mechanically producing interlinked structures
such as rotaxane and catenane [1516] without
using self-assembly (i.e., by adding hydrophobic ring structures to the molecular
parts which will then be attracted to each other in a polar solvent like water)
and thus allowing the positional assembly of very small (<1 kD) molecular
nanoscale parts.

Note that while most enzymes in cells are involved in manipulating
small molecules <0.25 kD, there are several classes of enzymes involved in
manufacturing complex covalently bound molecules such as vitamins, enzyme cofactors,
antibiotics, and toxins with masses up to ~3 kD. Molecules even larger than
this are manipulated by tRNA-synthetase (a 40-100 kD enzyme that manipulates
~30 kD tRNAs), the spliceosome, the ribosome, the proteosome, and the DNA replication
complex. (Many of these also involve “parts insertion” or “threading”
maneuvers, such as the clamp and bridge helix mechanisms in RNA polymerase II
that act as a translocation ratchet to feed DNA through the enzyme interior
in order to produce mRNA [2049].) By designing
synthetic enzymes consisting of synthetic amino acids, we can envision grabbing
molecular parts in a solution and then, as the enzyme folds, bringing them into
proper alignment and causing them to react – which might be called “nanopart
synthetases” or “protein-directed parts assembly.” Of course,
RNA-based ribozymes [2050-2054]
may prove better suited than proteins for some reactions, so we are not limited
to using enzymes to form the covalent bonds required in nanoparts. Directed
evolution in bacteria might be employed to create such specialty enzymes –
as has already been done, for example, in converting a disulfide reductase (disassembly)
enzyme into a disulfide-forming (assembly) enzyme [2055].

Ratchet-action protein-based molecular motors are well-known
in biology [2056]. Conformational cascades
of a special genetic variant of yeast cell prions have already been used to
self-assemble silver- and gold-particle-based nanowires [2057,
2058]. The GTPase dynamin mechanoenzyme
– which self-assembles into rings or spirals, wrapping around the necks
of budding vesicles and squeezing, pinching them off, during cellular endocytosis
– is well-known [2059-2063].
Amyloid fibril lamination can be exploited for nanotube self-assembly [2064],
and artificial self-assembled peptides can orient carbon nanotubes (CNTs) into
fibers [2065], achieve hydroxyapatite crystallization
[2066, 2067],
or grow copper nanocrystals [2068]. Smith
[1153, 2069]
has used methyltransferase-directed addressing of fusion proteins to DNA scaffolds
to construct a molecular camshaft (Figure
4.27) as a exemplar protein/nucleic acid biostructure. Bachand and Montemagno
[2070] have engineered a biomolecular motor
constructed of F1-ATPase protein [2071]
with an attached silicon nitride “propeller” arm [2072]
and a reversible on/off switch [2073],
and other task-optimized genetically engineered molecular motors have been synthesized
by others [2074]. Protein-protein binding
specificity has been used to bend silicon microcantilevers [2002,
2004, 2007].
Finally, molecular chaperones are a group of proteins that assists in the folding
of newly synthesized proteins or in the refolding of denatured proteins. Genetically
engineered chaperonin protein templates (heat shock proteins serving as chaperone
molecules) can direct the assembly of gold (1.4, 5, or 10 nm) and CdSe semiconductor
quantum dots (4.5 nm) into nanoscale arrays [2075].

Immunoglobulin (Ig) or antibody molecules could be used first
to recognize and bind to specific faces of crystalline nanoparts, then as handles
to allow attachment of the parts into arrays at known positions, or into even
more complex assemblies. As reported by Freitas [235]
(next 3 paragraphs): Kessler et al [2076]
raised monoclonal antibodies (MAbs) specific for crystals of 1,4-dinitrobenzene
having well-defined molecular-level structures. These antibodies were so specific
they would not bind to the same molecule when it was conjugated to a protein
carrier. IgG antibodies isolated from the serum of rabbits injected with crystals
of monosodium urate monohydrate or magnesium urate octahydrate evidently bear
in their binding sites an imprint of the crystal surface structure because they
can act as nucleating templates for crystal formation in vitro with extremely
low cross-reactivity, despite the similar molecular and structural characteristics
of the two crystals [2077]. Antibody binding
to monosodium urate crystals has been known for decades [2078],
and viruses have been engineered with a specific recognition moiety for ZnS
nanocrystals used as quantum dots [2160].
Interestingly, antibodies specific to in vivo water-ice crystals have even been
reported [2079].

Like antigens with ordered multiple epitopes (the antigenic
determinants), crystals expose chemically and geometrically distinct surfaces,
so different antibodies might recognize distinct faces of a crystal (possibly
including diamond or sapphire crystal faces) in an interaction similar to that
of antibodies for repetitive epitopes present on protein surfaces [2080,
2090]. For instance, one MAb to 1,4-dinitrobenzene
crystals was shown to specifically interact with the molecularly flat, aromatic,
and polar (101) face of these crystals, but not with other faces of the same
crystal [2088]. MAbs have also been elicited
against cholesterol monohydrate crystals [2081,
2082], one of which [2081]
was shown to specifically recognize the crystal’s stepped (301) face.
Here, the hydrophobic cholesterol hydrocarbon backbone is exposed on one side
of the molecular steps while hydroxyl residues and water molecules are exclusively
exposed on the other side. In both cases, crystal-specific antibodies were of
the IgM idiotype [2090]. This accords with
the assumption that (unlike most commonly used antigens) crystals cannot be
processed by the antigen presenting cells, hence antibodies must be induced
through a T cell-independent path [2083].
Semiconductor-binding [2160, 2163]
and calcite-binding [2084] proteins are
known that can discriminate among the various crystal faces of the given material
and can in some cases alter the pattern of crystal growth [2085].
Sulfur-free gold-binding proteins (GBPs) recognize and noncovalently bind preferentially
to the Au (111) crystal surface – GBPs use multiple repeats of 14-30 residue
sequences to bind to this surface [2086].
Hyaluronan is believed to be a crystal-binding protein for calcium oxalate monohydrate
crystals [2087].

Solubilized (derivatized) C60 and C70
fullerenes can induce the production of specific antibodies [2089-2093],
usually by interaction with the combining sites of IgG [2089].
It is speculated that highly hydrophobic pure fullerenes would be recognized
by antibodies with hydrophobic amino acids in their binding sites [2089,
2094] or would interact with donor -NH2
[2098] and -SH [2099]
groups. There are now many reports of antibodies being raised to single-walled
carbon nanotubes [2093-2097].
For example, antibodies raised to C60 in mice strongly bind to single-walled
nanotubes [2097]. Computer simulations
suggest that it may be possible to build antibodies that selectively bind to
nanotubes of a specific diameter or chirality [2094].